A culture method giving substantial yields of normal nasopharyngeal epithelial cells for work with Epstein-Barr virus

A culture method, utilising a feeder layer of lethally irradiated 3T3 fibroblasts and medium supplemented with hydrocortisone, cholera toxin, and epidermal growth factor, has been elaborated for the in vitro growth of normal human nasopharyngeal epithelial cells. This method allowed the cells to be grown in vitro for periods of up to 146 days, very considerably longer than in previously reported studies, and ensured that the cultures remained largely free from contaminating human fibroblasts. It was found possible to subculture the nasopharyngeal epithelial cells through numerous passages both by dispersing monolayers into single cell suspensions and by transferring coverslip monolayers of the cells to individual Petri dishes. By combining these two methods, at least 50 replicate epithelial cultures could be produced from each tissue sample, thus providing for the first time cultured nasopharyngeal epithelial cells in quantities suitable for extensive experiments with Epstein-Barr virus.